Table 4.

Comparison of advanced glycation end product detection techniques in bone tissue and their application limitations

Method	Minimum sample size	Spatial resolution	Absolute quantification	Major advantages	Limitations	Ref.
HPLC-FLD/LC-MS	2 mg defatted bone powder		Yes	High sensitivity; can differentiate CML/CEL/MG-H1/Pen	Destructive; requires acid hydrolysis; labor-intensive	[62]
Autofluorescence (Ex 335/Em 385)	5 μm tissue section	Micron level	No (relative)	Rapid, high-throughput; suitable for biopsy screening	Interference from mineral/Lipid autofluorescence; cannot distinguish AGE types	[63]
Raman spectroscopy	Applicable to both in vivo and tissue sections	Approximately 1 μm	No (semiquantitative)	In situ detection; simultaneously captures mineral-matrix information	Sensitive to water; spectrum interpretation requires expertise	[64]
Nano-FTIR/AFM-IR	10 μm tissue section	20-50 nm	No (semiquantitative)	Highest spatial resolution; enables single-fiber localization	Expensive equipment; limited scanning area	[65]

HPLC: High-performance liquid chromatography; FLD: Fluorescence detector; LC-MS: Liquid chromatography-mass spectrometry; Ex: Excitation; Em: Emission; Nano-FTIR: Nanoscale Fourier transform infrared spectroscopy; AFM-IR: Atomic force microscopy-infrared spectroscopy; CML: Nepsilon-(carboxymethyl)lysine; CEL: Nepsilon-(carboxyethyl)lysine; MG-H1: Methylglyoxal-derived hydroimidazolone 1; Pen: Pentosidine; AGE: Advanced glycation end-product.